Urquiza-Zurich, S. | bioRxiv Science

bioRxiv2025

SiteCELL enables on-site PBMCs purification and cryopreservation for immune single cell profiling of diverse ancestries

Single cell genomics has improved our knowledge of immune function and heterogeneity. In recent years, the steady increase in the number of cells and individuals profiled as part of large multinational projects has enabled the characterization of cellular differences across human populations and ancestries. However, methods for collecting and processing peripheral blood mononuclear cells (PBMCs) for downstream single cell sequencing are difficult to implement in remote and rural settings. This has resulted in a lack of representation of underserved communities across the Global South in current initiatives. Hence, we developed SiteCELL, a method that enables purification and cryopreservation of PBMCs from whole blood at the site of collection using minimal laboratory equipment and without electricity. By comparing matched samples of purified PBMCs, we showed that SiteCELL performs as well as ficoll density gradient (FDG), both in laboratory and rural settings. This method ensures accurate recovery of cell type proportions and excels in reducing stress and minimizing variability across sampling batches. These advantages make it particularly well suited for implementation in challenging settings across countries, thereby enabling the inclusion of underrepresented ancestries in cellular atlases.

immunology↗

bioRxiv2022

The assembly of wild natural isolates define neuronal integrity and life history traits of co-inhabiting C. elegans.

Bacterivore nematodes are the most abundant animals in the biosphere, largely contributing to global biogeochemistry. The effect of environmental microbes as source of associated microbiota and natural diet on their life history traits of nematodes is likely to impact the general health of the biosphere. Caenorhabditis elegans is a unique model to study the behavioral and physiological outputs of different available microbial diets. Nonetheless, most studies are on monoaxenic cultures of laboratory bacteria while the effect of natural microbiota isolates has only recently started to be reported. Here, we quantified physiological, phenotypical and behavioral traits of worms feeding on two bacteria that co-isolated with wild nematodes and tested how combinations of these isolates with other bacteria affected the traits measured. These bacteria were identified as a putative novel species of Stenotrophomonas denominated Stenotrophomonas sp. Iso1 and a strain of Bacillus pumilus designated Iso2. The isolates induced distinctive behaviors and development patterns that changed in mixes of the two bacteria and/or the pathogen Salmonella enterica. Focusing on the degeneration rate of the touch circuit of C. elegans we show that B. pumilus alone is protective while the mix with Stenotrophomonas sp. is degenerative. The analysis of the metabolite content of each isolate and their combination identified NAD+ as potentially neuroprotective. In vivo supplementation shows that NAD+ restores neuroprotection to the mixes and also to individual non-protective bacteria. The results highlight the need to study the physiological effects of bacteria resembling native diets in a multicomponent scenario rather than using single isolates. ImportanceThe behavioral decisions of animals depend on their microbiota. In nature it is unknown how this interaction affects the health of the biosphere. To study how the nematode-bacteria relationship impacts the life history traits of these animals, we isolated bacteria found in association with wild nematodes and tested their influence as single species and consortia, in the life history traits of the model C. elegans. We identify metabolites from wild bacteria that change these traits. The bacteria isolated were identified a Stenotrophomonas sp and a B. pumilus. We find that all traits depend on the biota composition. For example, B. pumilus is neuroprotective to degenerating neurons of the touch circuit of C. elegans needed to sense and escape from predators in the wild. The co-culture with Stenotrophomonas sp. eliminates the protection. We identified NAD+ as the metabolite lost in the mix, and show that NAD+ by itself is neuroprotective.

microbiology↗